The effect of 475°C embrittlement on microstructure development of grade 2205 duplex stainless steel was investigated. Spinodal decomposition products and associated precipitates in ferrite, austenite, and at interphase boundaries were characterized using analytical transmission electron microscopy (TEM) and scanning electron microscopy (SEM) techniques. Micro-analyses confirmed the presence of Cr-enriched α'' and Cr-depleted α' spinodal structures in the ferrite after 5 h of aging at 475°C. Long-term aging for 255 h resulted in heavily-faulted R-phase precipitates with sizes of ∼50–400 nm, χ-phase, and ε-Cu in the ferrite, TiN and Cr2N precipitates in the austenite, and a continuous network of M23C6-carbides at interphase boundaries. A significant hardness increase was observed after 255 h of aging, which was accompanied by a reduction of ferrite fraction. X-ray diffraction (XRD) stress measurements showed a general reduction of residual stresses in both ferrite and austenite with aging. Electron backscatter diffraction (EBSD) showed increased local misorientations, primarily close to precipitate interfaces within the ferrite, indicating the development of strain heterogeneities in the microstructure. The data presented provided a better understanding of 475°C embrittlement in duplex stainless steel, suggesting that not only the ferrite alone is responsible for embrittlement. A comprehensive microstructure characterization study has been provided and the explanation for 475°C embrittlement of duplex stainless steel has been discussed.